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Biomechanical remodeling of stroma by cancer-associated fibroblasts (CAF) in early stages of cancer is critical for cancer progression, and mechanical cues such as extracellular matrix stiffness control cell differentiation and malignant progression. However, the mechanism by which CAF activation occurs in low stiffness stroma in early stages of cancer is unclear. Here, we investigated the molecular mechanism underlying CAF regulation by SPIN90 and microtubule acetylation under conditions of mechanically soft matrices corresponding to normal stromal rigidity. SPIN90 was downregulated in breast cancer stroma but not tumor, and this low stromal expression correlated with decreased survival in breast cancer patients. Spin90 deficiency facilitated recruitment of mDia2 and APC complex to microtubules, resulting in increased microtubule acetylation. This increased acetylation promoted nuclear localization of YAP, which upregulated expression of myofibroblast marker genes on soft matrices. Spin90 depletion enhanced tumor progression, and blockade of microtubule acetylation in CAF significantly inhibited tumor growth in mice. Together, our data demonstrate that loss of SPIN90-mediated microtubule acetylation is a key step in CAF activation in low stiffness stroma. Moreover, correlation among these factors in human breast cancer tissue supports the clinical relevance of SPIN90 and microtubule acetylation in tumor development. Cancer Res; 77(17); 4710–22. ©2017 AACR.

Supplementary Table S2 shows the clinicopathological characteristics of clinical samples.

Figure S1 (Related to Fig. 1). Low expression of SPIN90 in cancer stroma and CAFs. Figure S2 (Related to Fig. 2). CAFs derived from Spin90-/- mice show increased activation. Figure S3 (Related to Fig. 3). Loss of SPIN90 induces CAF differentiation. Figure S4 (Related to Fig. 4). Recruitment of mDia2 to acetylated microtubules regulates CAF differentiation. Figure S5 (Related to Fig. 5). Acetylated α-tubulin enhances nuclear translocation of YAP on 0.5 kPa PAGs. Figure S6 (Related to Fig. 6). Increased α-tubulin acetylation in Spin90-/- mice induces cancer progression. Table S1. Primers used for RT-PCR

Supplementary methods and References - Supplementary methods describing key reagents, plasmids, cell culture, CRISP/Cas9 system, HER2 agonist preparation, RT-PCR, western blot, flow-cytometry, tumorsphere culture, ChIP assay, animal experiments, Immunohistochemically staining and microarray analysis.

Systemic inflammation in breast cancer correlates with poor prognosis, but the molecular underpinnings of this connection are not well understood. In this study, we explored the relationship between HER2 overexpression, inflammation, and expansion of the mammary stem/progenitor and cancer stem–like cell (CSC) population in breast cancer. HER2-positive epithelial cells initiated and sustained an inflammatory milieu needed to promote tumorigenesis. HER2 induced a feedforward activation loop of IL1α and IL6 that stimulated NFκB and STAT3 pathways for generation and maintenance of breast CSC. In mice, Il1a genetic deficiency delayed MMTV-Her2–induced tumorigenesis and reduced inflammatory cytokine expression as well as CSC in primary tumors. In clinical specimens of human breast tumor tissues, tissue microarray analysis revealed a strong positive correlation between IL1α/IL6 expression and CSC-positive phenotype. Pharmacologic blockade of IL1α signaling reduced the CSC population and improved chemotherapeutic efficacy. Our findings suggest new therapeutic or prevention strategies for HER2-positive breast cancers.Significance: IL1α signaling driven by HER2 promotes chronic inflammation needed to support cancer stem-like cell maintenance in HER2-positive breast cancers. Cancer Res; 78(8); 2040–51. ©2018 AACR.

Supplementary Tables - Supplementary Tables S1, S3 and S4. S1 shows frequency of CSCs after transplantation of the indicated cells. S3 shows multivariate Cox regression data. S4 Positive correlation between IL-1α and HER2 or IL6 expression in breast cancer tissues.

Background: Human epididymis protein 4 (HE4), also known as WFDC2, is a member of whey acidic protein (WAP) family with a presumptive role in natural immunity. HE4 expression has been increased in many cancer types, particularly in gynecologic and pulmonary cancers. HE4 has been identified as an important serum biomarker in the diagnosis and monitoring of epithelial ovarian cancer. Recent researches have demonstrated that HE4 levels in serum and tissues are elevated in patients with breast cancer and HE4 is thought be involved in breast cancer progression. In contrast, expression of HE4 in ductal carcinoma in situ (DCIS) has not been well characterized yet. The present work was undertaken to evaluate serum and tissue levels of HE4 in DCIS and their correlations with clinicopathological features of DCIS. Materials and methods: Preoperative serum HE4 levels in 59 DCIS patients was analyzed using the ARCHITECT assay. Tissue microarray containing DCIS and adjacent normal tissues from the same cohort were tested for HE4 mRNA and protein by RNAscope in situ hybridization (ISH) and immunohistochemistry analysis, respectively. Tissue microarray containing only DCS tissues from 41 patients were also tested for HE4 mRNA and protein. To validate prognostic potential of HE4 in breast cancer patients, the BreastMark database was used. Results: HE4 levels in 59 DCIS patients ranged from 23.5 to 86.3 pmol/L (mean ± SD: 39.4 ± 11.9). Based on the cutoff level, there were no abnormal cases for HE4. Serum HE4 levels did not differ according to clinicopathological parameters except for menopause status. RNAscope ISH and immunohistochemistry confirmed an increase in HE4 in DCIS tissues compared with their corresponding normal tissues. Spearman’s correlation analyses revealed no significant correlation between serum and tissue levels of HE4. Among 100 DCIS tissues, there was a positive correlation between HE4 mRNA ISH scores and HE4 immunohistochemical staining scores (r = 0.771, P < 0.001). High mRNA and protein expression of HE4 were observed in 25 of 99 (25.3%) and 34 of 99 (34.3%) cases, respectively. High mRNA HE4 expression was significantly associated with low stromal tumor infiltrating lymphocyte (TIL) density scores, ER positivity, HER2 negativity, and HR+/HER2- subtype. High protein HE4 expression was also significantly associated with absence of comedo-type necrosis, low stromal TIL density scores, ER positivity, HER2 negativity, and HR+/HER2- subtype. HE4 mRNA and protein expression did not correlate with recurrence of DCIS. In breast cancer patients, high HE4 expression was significantly associated with good survival in the overall group (HR = 0.838, P = 0.003, n = 2,652) and lymph node negative group (HR = 0.791, P = 0.029, n = 1,183). Conclusions: Our study revealed that serum HE4 is not elevated in patients with DCIS. High expression of HE4 mRNA and protein in DCIS tissues are associated with good clinicopathological characteristics, which warrant. further investigations. Citation Format: Ji Shin Leee, Nah Ihm Kim, Min Ho Park. Evaluation of human epididymis protein 4 serum and tissue expression in ductal carcinoma in situ of the breast [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P4-07-22.

Purpose: Pathologic N3a breast cancer is defined as having 10 or more metastatic axillary lymph nodes and metastases to the infraclavicular (level III axillary lymph) node. This study aimed to determine clinicopathological factors and assess the importance of lymph node ratio (LNR) as a prognostic factor for patient with N3a breast cancer treated with surgery without neoadjuvant chemotherapy.Methods: Medical records of 154 patients who underwent surgery were retrospectively analyzed. Patients were pathologically diagnosed with N3a breast cancer between May 2004 and December 2014. LNR was defined as the number of metastatic lymph nodes divided by the total number of resected lymph nodes. It was calculated using the receiver operating characteristic (ROC) curve. The median follow up period was 94 months (range, 10–205 months).Results: Among 154 patients with N3a breast cancer, 70 (45.5%) had recurrence and 40 (26.0%) died during the follow-up period. The 5-year disease free survival (DFS) and overall survival (OS) rates after surgery were 63.0% and 85.9%, respectively. LNR>0.82 (hazard ratio [HR]: 2.271; 95% confidence interval [CI]: 1.413–3.649; p=0.001) was a prognostic factor significantly associated with DFS. LNR>0.68 (HR: 2.156; 95% CI: 1.146–4.044; p=0.017) and invasive ductal carcinoma (HR: 0.125; 95% CI: 0.017–0.915: p=0.041) were significantly associated with OS.Conclusion: Although LNR values associated with DFS and OS are slightly different, LNR is a good prognostic factor for patients with N3a breast cancer.

Adenoma of the nipple is a rare benign type of breast that occurs most often in middle-aged women with nipple discharge, skin erosion in addition to crusting, inflammation, and pain. It can be clinically confused with Paget’s disease or breast cancer precursor lesions, such as ductal carcinoma in situ low grade. The treatment of choice for nipple adenoma is complete excision of the tumor. However in younger women, nipple preservation is required. We present two cases of nipple adenoma that were completely removed with nipple preservation.

Purpose: Intraoperative frozen section analysis to assess margin status during breast-conserving surgery is often performed to reduce the rate of re-excision. Whether additional resection is required if atypical cells are found at the margin during breast-conserving surgery is controversial. The aim of this study was to evaluate the accuracy of intraoperative frozen section analysis and investigate the feasibility of additional resection in cases of atypical hyperplasia on frozen sections from breast-conserving surgery.Methods: A retrospective analysis was performed on 1,411 patients with invasive breast cancer who underwent breast-conserving surgery between July 2004 and June 2012. The microscopic margins of the intraoperative frozen sections and permanent sections were examined. Overall events (locoregional recurrence and distant metastasis) were analyzed between the negative margin group and the other margin group (including atypical hyperplasia, carcinoma in situ [CIS], and invasive carcinoma).Results: Of the 1,411 patients, 1,201 had negative margins and 210 had other margin types in the first frozen section. 68 patients had atypical hyperplasia on frozen section analysis. This atypical hyperplasia included atypical cells, atypical ductal hyperplasia, and atypical lobular hyperplasia. Of these 68 patients, the final findings on the permanent section were negative (n=32, 47.1%), atypical cells (n=16, 23.5%), and CIS (n=20, 29.4%). The rate of overall events and distant metastases were higher in the other margin group (9.5% vs. 5.4%, p=0.021) and (5.7% vs. 3.0%, p=0.045). Of the 1,411 patients, 44 (3.1%) had false-positive findings, and 15 (1.1%) had false-negative findings in the frozen sections, with an accuracy of 95.8%.Conclusion: The accuracy of intraoperative frozen section analysis was 95.8%, with 91.7% sensitivity and 96.4% specificity. Atypical hyperplasia on frozen section analysis requires additional resection at the time of breast-conserving surgery because of the possibility of CIS, but excessive resection may be performed in other cases. Therefore, sufficient consultation with patients is required.

BACKGROUND/AIM Thyroid lobectomy may cause post-lobectomy hypothyroidism. We investigated the difference in levothyroxine (LT4) supplementation and cessation between patients with benign disease and those with papillary thyroid carcinoma (PTC) and found that the rate of LT4 cessation could be decreased after thyroid-stimulating hormone (TSH) suppression in PTC. PATIENTS AND METHODS We retrospectively reviewed 88 patients with benign tumor and 463 patients with PTC and investigated the risk factors for LT4 supplementation after thyroid lobectomy. RESULTS During the median follow-up of 73.0 months, 207 (37.6%) patients maintained the euthyroid state, while 344 (62.4%) patients continued LT4 supplementation for LT4 replacement or TSH suppression. In patients with benign tumors, only high pre-TSH level (>1.98 mIU/l) was a significant risk factor (odds ratio [OR]=10.09). However, in patients with PTC, pre-TSH level ≥1.98 mIU/l (OR=3.28), pregnancy planning (OR=2.97), and age ≥42.5 years (OR=1.94) were significant risk factors. Moreover, the most potent risk factor was tumor aggressiveness (OR=4.00), which was found to be more significant than high pre-TSH. The overall rate of LT4 cessation in all patients was 37.6%; however, in the 303 patients who underwent the LT4-Off trial, there was no difference in the rate in the benign tumor, low-risk PTC, and intermediate-risk PTC groups (66.2%, 68.8%, and 70.8%, respectively; p=0.886). CONCLUSION When post-lobectomy TSH levels were adequate and the risk of recurrence was reduced, LT4 cessation in PTC could be achieved at the same rate as that in benign tumors, regardless of the duration of TSH suppression.

One of the major obstacles in the way of high-performance quantum dot light-emitting diodes (QLEDs) is the charge imbalance arising from more efficient electron injection into the emission layer than the hole injection. In previous studies, a balanced charge injection was often achieved by lowering the electron injection efficiency; however, high performance next-generation QLEDs require the hole injection efficiency to be enhanced to the level of electron injection efficiency. Here, we introduce a solution-processed HfO

By combining data from 160,500 individuals with breast cancer and 226,196 controls of Asian and European ancestry, we conducted genome- and transcriptome-wide association studies of breast cancer. We identified 222 genetic risk loci and 137 genes that were associated with breast cancer risk at a p

Parkinson’s disease (PD), characterized by degeneration of dopaminergic neurons, share pathogenic features with obesity, including mitochondrial dysfunction and oxidative stress. Paraoxonase 2 (PON2) is an inner mitochondrial membrane protein that is highly expressed in dopaminergic neurons and is involved in the regulation of mitochondrial oxidative stress. However, no drug targeting PON2 has ever been developed for the treatment of PD. Here, we show that vutiglabridin, a clinical phase 2-stage drug for the treatment of obesity, has therapeutic effects in PD models, targeting mitochondrial PON2. Vutiglabridin penetrates into the brain, binds to PON2, and restores 1-methyl-4-phenylpyridinium (MPP+)-induced mitochondrial dysfunction in SH-SY5Y neuroblastoma cells. Knockdown of PON2 by lentiviral shRNA infection abolished the effects of vutiglabridin on mitochondria. In mice, vutiglabridin significantly alleviated motor impairments and damage to dopaminergic neurons in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD model, and these effects were also abolished in PON2-knockdown mice, suggesting that vutiglabridin is neuroprotective via PON2. Extensive in vitro and in vivo assessment of potential neurotoxicity showed vutiglabridin to be safe. Overall, these findings provide support for the clinical development of vutiglabridin as a novel PON2 modulator for the treatment of PD.One Sentence SummaryTargeting paraoxonase-2 by a clinical-stage compound vutiglabridin provides neuroprotective effects in preclinical models of Parkinson’s disease.

In recent years, postponing childbearing has increased the prevalence of pregnancy-associated breast cancer (PABC). PABC has a poorer prognosis than breast cancer not associated with pregnancy (non-PABC) due to delayed diagnosis and aggressive subtype. Additionally, pregnancy itself predicts a poor prognosis; but, this is a subject of debate. Thus, we analyzed the effects of known prognostic factors and pregnancy on the prognosis of PABC. We retrospectively analyzed women aged 20 to 49 years who were diagnosed with breast cancer (BC) between 1989 and 2014. Patients were distributed into PABC and non-PABC groups, and 1:4 propensity score matching was performed to adjust for baseline characteristics. Primary endpoints were overall survival (OS) and BC-specific survival (BCSS). Secondary endpoint was the difference in prognosis according to BC subtype. Of the 34,970 recruited patients with BC, 410 (1.2%) had PABC. Patients with PABC were younger and tended to have triple-negative BC (TNBC) subtype than non-PABC patients. The 1640 matched non-PABC patients showed a significantly worse mean survival rate than the unmatched non-PABC patients. Patients with PABC had a significantly worse OS and BCSS than those with non-PABC. In multivariate analyses, patients with PABC of luminal B (Ki-67 ≥14.0%) and TNBC subtypes had worse OS and BCSS than patients with non-PABC. Patients with PABC had poorer prognosis than non-PABC patients after adjusting for several prognostic factors. This difference was particularly significant in patients with the luminal B and TNBC subtypes.

Eco-friendliness is an important global issue, and the maritime field is no exception. Predicting the composition of exhaust gases emitted by ship engines will be of consequence in this respect. Therefore, in this study, exhaust gas data were collected from the generator engine of a real ship along with engine-related data to predict emission characteristics. This is because installing an emission gas analyzer on a ship has substantial economic burden, and, even if it is installed, the accuracy can be increased by a virtual sensor. Furthermore, data were obtained with and without operating the SCR (often mounted on ships to reduce NOx), which is a crucial facility to satisfy environment regulation. In this study, four types of datasets were created by adding cooling and electrical-related variables to the basic engine dataset to check whether it improves model performance or not; each of these datasets consisted of 15 to 26 variables as inputs. CO2 (%), NOx (ppm), and tEx (°C) were predicted from each dataset using an artificial neural network (ANN) model and a support vector machine (SVM) model with optimal hyperparameters selected by trial and error. The results confirmed that the SVM model performed better on smaller datasets, such as the one used in this study compared to the ANN model. Moreover, the dataset type, DaCE, which had both cooling and electrical-related variables added to the basic engine dataset, yielded the best overall prediction performance. When the performance of the SVM model was measured using the test data of a DaCE on both no-SCR mode and SCR mode, the RMSE (R2) of CO2 was between 0.1137% (0.8119) and 0.0912% (0.8975), the RMSE (R2) of NOx was between 17.1088 ppm (0.9643) and 13.6775 ppm (0.9776), and the RMSE (R2) of tEx was between 4.5839 °C (0.8754) and 1.5688 °C (0.9392).

Although adjuvant chemotherapy (CTx) is still recommended for high-risk patients with hormone receptor-positive and human epidermal receptor (HER)-2-negative breast cancer, recent studies found that selected patients with low disease burden may be spared from CTx and receive hormonal treatment (HT) alone. This study aims to evaluate the trends of treatment (CTx + HTThe Korean Breast Cancer Society Registry was queried (2000 to 2018) for women with pT1-2N0-1 hormone receptor-positive and HER2-negative disease who underwent surgery and adjuvant systemic treatment (CTx and HT). Clinicopathologic factors, change in pattern of treatment over time, and OS for each treatment option were analyzed.A total of 40,938 women were included in the study; 20,880 (51.0%) received CTx + HT, while 20,058 (49.0%) received HT only. In recent years, there has been a steady increase in the use of HT alone, from 21.0% (2000) to 64.6% (2018). In Cox regression analysis, age, type of breast and axillary operations, T and N stages, body mass index, histologic grade, and presence of lymphovascular invasion were prognostic indicators for OS. There was no significant difference between CTx + HT and HT alone in terms of OS (P = 0.126).Over the years, there has been a shift from CTx + HT to HT alone without a significant difference in OS. Therefore, HT alone could be a safe treatment option in selected patients, even those with T2N1 disease.

Activation of mutations in RAS has been found in high ratio in human cancers, inducing downstream critical effectors corresponding tumorigenesis. In recent years, RAS-RAF-ERK1/2 pathway is known as a crucial target for the anti-cancer drug development because of high prevalence of ERK activation in human cancers. The attempts in direct targeting to activated RAS, which is GTP bound, has been somewhat successful in the treatment of certain type of cancer, however, still beset by the resistance. Our aim is to develop inhibitor that cell-penetrating scFv (single-chain variable fragment) which targets mutated KRAS (mKBscFv; mutated KRAS binding scFv). The Fv fragment is the smallest unit composed of the variable region of heavy chain and light chain in an immunoglobulin molecule, that has a function of antigen-binding activity. The scFv can specifically bind to an antigen, thereby reducing side effects of non-specific binding seen in chemical drugs. However, due to the lack of cell penetrating ability of scFv, a delivery carrier is often required. In this study, the mKBscFv was measured binding affinity toward wild type KRAS and mutated KRAS using surface plasmon resonance (SPR). The scFv has higher affinity to mutated KRAS than wild type KRAS. A cell-penetrating peptide (CPP) was developed for drug delivery and was expressed at the C-terminus of mKBscFv for transferring cells. The mKBscFv-CPP was tagged with His tag to express in CHO cells, which was purified by FPLC. The purity of mKBscFv-CPP was confirmed by SDS PAGE and immunoblotting. The mKBscFv-CPP treated KRAS mutated cancer cells decreased cell viability in a concentration-dependent manner, and decreased expression of mutated GTP-bound KRAS. In addition to the bioactivity of the mKBscFv, it has further advantage as a proteolysis-targeting chimera (PROTAC). The plasmid DNA was prepared by adding a VHL-expressing sequence to the C-terminus of mKBscFv (mKBscFv-Vh). This gene was transfected into KRAS mutated cancer cells to express the protein, and as a result mutated GTP-bound KRAS was reduced. A CPP peptide domain was added to the C-terminus of this gene to express it as a protein in CHO cells. The mutated GTP-bound KRAS was reduced by mKBscFv-Vh-CPP in KRAS mutated cacner cells. Taken together, mKBscFv that binds to mutated KRAS was developed, and it can be expected to become a new protein anticancer drug effective against mutated KRAS-mediated cancer cells by fusing cell penetrating peptide and PROTAC technology. Citation Format: Gookjin Yoon, Beom Soo Jo, Dong Woo Lee, Jinwook Yang, Min-Ho Park, Sanghui Seok, Jue Yeon Lee, Chong Pyoung Chung, Yoon Shin Park, Yoon Jeong Park. BIO-PROTAC: Application of cell penetrating scFv to treat undruggable KRAS mutant cancer [abstract]. In: Proceedings of the AACR Special Conference: Targeting RAS; 2023 Mar 5-8; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Res 2023;21(5_Suppl):Abstract nr B039.

Activation of mutations in RAS has been found in high ratio in human cancers, inducing downstream critical effectors corresponding tumorigenesis. The attempts in direct targeting to activated RAS, which is GTP bound, has been somewhat successful in the treatment of certain type of cancer, however, still beset by the resistance. It has been well established that the tumor engagement is quite related with the interaction between mutated RAS protein and RAF kinase in ERK cascade. Our aim is to develop inhibitor that cell-penetrating scFv (single chain fragment variable) which targets mutated KRAS (mKBscFv). The Fv fragment is the smallest unit composed of the variable region of heavy chain and light chain in an immunoglobulin molecule, that has a function of antigen-binding activity. The scFv can specifically bind to an antigen, thereby reducing side effects of non-specific binding seen in chemical drugs. However, due to the lack of cell penetrating ability of scFv, a delivery carrier is often required. In this study, a cell-penetrating synthetic peptide (CPP) was developed for drug delivery carrier and was expressed at the C-terminus of mKBscFv for transferred cells. In addition, the CPP peptide has a function of inhibiting the interaction of mutated RAS and RAF. The mKBscFv-CPP specifically binding to mutated KRAS and tagged with His tag to express in CHO cells, which was purified by FPLC. The purity of mKBscFv-CPP was confirmed by SDS PAGE. In the SPR results, not only mKBscFv-CPP showed high binding ability to mutated KRAS, but also in cell viability test, it affected the proliferation of KRAS mutated cells. The muatated GTP-bound KRAS was reduced by mKBscFv in KRAS mutated cancer cells. In addition to the bioactivity of the mKBscFv, it has further advantage as a proteolysis-targeting chimera (PROTAC). The plasmid DNA was prepared by adding a VHL-expressing sequence to the C-terminus of mKBscFv (mKBscFv-Vh). This gene was transfected into KRAS mutated cancer cells to express the protein, and as a result mutated GTP-bound KRAS was reduced. A cell penetrating synthetic peptide domain was added to the C-terminus of this gene to express it as a protein in CHO cells. The mutated GTP-bound KRAS was reduced by mKBscFv-Vh-CPP in KRAS mutated cancer cells. Taken together, mKBscFv that binds to mutated KRAS was developed, and it can be expected to become a new protein anticancer drug effective against mutated KRAS-mediated cancer cells by fusing cell penetrating peptide and PROTAC technology. Citation Format: Gookjin Yoon, Beom Soo Jo, Dong Woo Lee, Jinwook Yang, Min-Ho Park, Sanghui Seok, Jue-Yeon Lee, Chong Pyung Chung, Yoon Jeong Park. BIO-PROTAC: New treatment modality to treat undruggable KRAS mutants by application of cell penetrating scFv [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 569.

Background: Triple-negative breast cancer (TNBC) is a heterogeneous group of cancer with dismal prognosis. B7 families can be promising targets for cancer immunotherapy in TNBC. Previously we have found increased expression of B7-H3 and B7-H4 mRNA and protein in breast cancer including TNBC. In an effort to discover therapeutic targets, TNBC has been further stratified into molecular subtypes. However, little is known about the clinical impact and value of B7-H3 and B7-H4 in TNBC subtypes. The purpose of this study was to evaluate the clinicopathologic characteristics of the B7-H3 and B7-H4 mRNA and protein expression according to the TNBC subset. Materials and methods: A tissue microarray was constructed from 186 patients with TNBC. B7-H3 and B7-H4 mRNA and protein expression were assessed by the RNAscope in situ hybridization (ISH) and immunohistochemistry. Immunohistochemistry for the TNBC molecular subtype-surrogate markers [cytokeratin 5/6 (CK5/6), CK14, epidermal growth factor receptor (EGFR), and androgen receptor (AR)], CD3, and CD8 was also performed. TNBC subtypes were classified into three subtypes: basal-like (BL), luminal AR (LAR), and unclassifiable type (UN). Results: Based on the immunohistochemical results, 186 TNBCs were classified into BL (n=120, 64.5%), LAR (n=10, 10.8%), and UN (n=46, 24.7%) subtypes. BL and UN subtypes were associated with younger age than the LAR subtype. B7-H3 mRNA and protein expressions were expressed in the tumor and stromal cells of the TNBCs. On the contrary, B7-H4 mRNA and protein expressions were only observed in the tumor cells. High tumor mRNA and protein expression of B7-H3 and B7-H4 were found in 49 of 175 (28.0%) and 121 of 176 (68.8) cases, and 92 of 174 (52.9%) and 66 of 176 (37.5%) cases, respectively. High stromal B7-H3 mRNA and protein expression were observed in 22 of 175 (12.6%) and 43 of 176 (24.4%) cases. High stromal B7-H4 mRNA and protein expression were not observed. B7-H3 and B7-H4 protein expression were closely correlated with their mRNA expression according to the tumor compartment. Tumor B7-H4 mRNA expression was associated with younger age at the initial diagnosis and molecular TNBC subtypes. Expression of B7-H3 mRNA and protein in the tumor cells negatively correlated with CD3+ and CD8+ T cell infiltration density in the tumor and/or stromal compartment of the TNBCs and its subtypes. High stromal B7-H3 mRNA expression was associated with poor disease-free and overall survival in the TNBCs and overall survival in the UN subtype. Stromal B7-H3 mRNA expression was independently associated with overall survival in the TNBCs and disease-free survival in the BL subtype. Conclusions: As prognostic factors, our results point to the importance of the expression of B7-H3 mRNA by the stromal cells in the TNBCs and the BL subtype. The inverse relationship between B7-H3 expression and CD3+ and CD8+ T lymphocyte infiltration may represent a promising target in the immunotherapy of the TNBCs, irrespective of its molecular subtypes. Citation Format: Ji Shin Lee, Nah Ihm Kim, Min Ho Park. B7-H3 and B7-H4 expression in triple-negative breast cancer subtypes detected by RNA in situ hybridization and immunohistochemistry: Association with clinicopathological features and T-cell infiltration [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS17-35.

Triple-negative breast cancer (TNBC) is relatively more aggressive than other subtypes of breast cancer. Lymphovascular invasion (LVI) is a well-known predictive factor of worse survival outcomes; however, the role of LVI in TNBC remains unclear. This study aimed to evaluate the significance of LVI and examine clinical outcomes in patients with TNBC. We reviewed the medical records of 417 patients with pathology results negative for estrogen receptor, progesterone receptor, and human epidermal growth factor receptor type 2 (immunohistochemical stain score 0, 1, or 2 with negative for fluorescence (or silver) in situ hybridization). Cox proportional hazards analysis was used to investigate the relationship between clinicopathological variables and survival outcomes. During a median follow-up of 93.8 months (range, 7.1−193.8), 65 (15.6%) patients had recurrence, and 37 (8.9%) patients died from cancer progression. In multivariate analyses of survival outcomes, the N1–3 stage (vs. N0 stage; hazard ratio [HR], 2.127; 95% confidence interval [CI], 1.237−3.659; p = 0.006), and LVI (vs. no LVI; HR, 2.053; 95% CI, 1.199−3.515; p = 0.009) were associated with worse recurrence-free survival. The N1–3 stage (vs. N0 stage; HR, 4.386; 95% CI, 2.140−8.988; p < 0.001) was associated with worse disease-specific survival. LVI in TNBC is an important predictive factor for recurrence than cancer-related death after adjustment for other prognostic factors. We suggest that patients with TNBC with LVI should receive short-term follow-up with imaging studies for detecting loco-regional and distant recurrence.

Background: Triple-negative breast cancer (TNBC) is a heterogeneous group of cancer with dismal prognosis. In an effort to discover therapeutic targets, TNBC has been further stratified into molecular subtypes. Immune checkpoint inhibitors targeting programmed cell death protein 1 (PD-1) and its ligand (PD-L1) have been increasingly utilized for the treatment of TNBC. Difficulty remains, however, in appropriate patient selection for treatment, as many PD-L1-positive cancers fail to show durable responses to PD-1/PD-L1 inhibition. Major histocompatibility complex (MHC) class I molecules play a crucial role in the presentation of tumor antigenic peptides to cytotoxic T cells. Loss of MHC class I expression in tumor cells represents a possible mechanism of immunotherapeutic resistance in PD-L1-positive cancers. However, little is known about the clinical impact and value of MHC class I expression in TNBC subtypes. Thus, this study examined MHC class I expression in TNBC subtypes with attention to PD-L1 expression and T cell infiltration. Materials and methods: MHC class I and PD-L1 expression were assessed by immunohistochemistry in 256 TNBC samples using tissue microarray. Immunohistochemistry for the TNBC molecular subtype-surrogate markers [cytokeratin 5/6 (CK5/6), CK14, epidermal growth factor receptor (EGFR), and androgen receptor (AR)], CD3, and CD8 was also performed. TNBC subtypes were classified into three subtypes: basal-like (BL), luminal AR (LAR), and unclassifiable type (UN). Results: All immunohistochemical markers were interpretable in 240 cases. TNBCs were classified into BL (65.8%), LAR (11.3%), and UN (22.9%) subtypes. Loss of MHC class I expression was found in 62 of 240 (25.8%) cases and was observed in 23.4% of BL, 40.7% of LAR, and 25.9% of UN subtype. PD-L1 expression in tumor-associated immune cells (≥1%) was seen in 69.2% (166/240). Among 166 PD-L1-positive TNBC cases, loss of MHC class I expression was seen in 16.4% of BL, 31.3% of LAR, and 20.0% of UN subtype. Loss of MHC class I expression was associated with low stromal TILs density and low CD3+ and CD8+ T-cell infiltration in BL subtype. CD3+ and CD8+ T-cell infiltration in TNBC subtype had a positive correlation with PD-L1 expression. In PD-L1 positive BL subtype, MHC class I lost expression showed a lower infiltration with CD3+ and CD8+ T-cells than MHC class I intact expression. In BL subtype, PD-L1 expression was associated with better disease-free survival and loss of MHC class I expression was associated with poor overall survival. However, MHC class I and PD-L1 expressions were not independent prognostic factors for disease-free or overall survival. Conclusions: Loss of MHC class I expression was found in TNBC, including PD-L1 positive cases. In PD-L1 positive BL subtype, loss of MHC class I expression was associated with low CD3 and CD8+ T-cell infiltration. Our results suggest that the evaluation of PD-L1 and MHC class I expression together is very important for the selection of potential responders to anti-PD-1/PD-L1 therapy for the TNBCs, especially the BL subtype. Citation Format: ji shin lee, nah ihm kim, min ho park. MHC class I and PD-L1 expression in triple-negative breast cancer subtypes: impact on T-cell infiltration and clinical outcome [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD9-02.

Purpose Approval of eribulin for metastatic breast cancer was based on data primarily from Western patients, and there is a paucity of data on the effectiveness and safety of eribulin for Asian patients. To determine the effectiveness and safety of eribulin in Korean women with breast cancer in a real-world setting, we conducted a nationwide, multicenter, retrospective study. Methods Patients with locally advanced or metastatic breast cancer who were treated with eribulin in 14 centers throughout Korea were included in this study. Eribulin was generally administered at a dose of 1.23 mg/m2 (equivalent to 1.4 mg/m2 eribulin mesylate) by intravenous infusion for 2–5 min, or as a diluted solution, on Days 1 and 8 of every 21-day cycle. The primary endpoint was progression-free survival (PFS) rate at 6 months. Secondary endpoints included median PFS, overall survival (OS), time-to-treatment failure (TTF), tumor response rate, and incidence of hematologic treatment-emergent adverse events (TEAEs). Results The safety and full analysis populations included 398 and 360 (38 had no efficacy data) patients, respectively. The PFS rate at 6 months was 37.8%. Median PFS, OS, and TTF were 134, 631, and 120 days, respectively. Objective response rate, clinical benefit rate, and disease control rate were 18.1%, 50.6%, and 49.4%, respectively. Hematologic TEAEs were reported in 65.1% of patients; neutropenia (56.8%) and anemia (11.3%) were most common. Conclusion Real-world effectiveness and safety of eribulin in Korean breast cancer patients were consistent with previous reports; no new safety concerns were identified., Graphical abstract Image 1, Highlights • Metastatic breast cancer patients were treated with eribulin (1.23 mg/m2, IV). • Progression-free survival rate at 6 months was 37.8% in eribulin-treated patients. • Median progression-free and overall survivals were 134 and 631 days, respectively. • Hematologic treatment-emergent adverse events were reported in 65.1% of patients. • Effectiveness and safety of eribulin were consistent with previous reports.

Skip lymph node (LN) metastases in papillary thyroid carcinoma (PTC) belong to N1b classification in the absence of central neck LN involvement. This study aimed to evaluate the predictive factors of skip metastases and their impact on recurrence in PTC patients with pN1b. A total of 334 PTC patients who underwent total thyroidectomy with LN dissection (central and lateral neck compartment) followed by radioactive iodine ablation were included. Patients with skip metastases tended to have a small primary tumor (≤1 cm) and single lateral neck level involvement. Tumor size ≤ 1 cm was an important predictive factor for skip metastases. Univariate analysis for recurrence showed that patients with a central LN ratio > 0.68, lateral LN ratio > 0.21, and stimulated thyroglobulin (Tg) levels > 7.3 ng/mL had shorter RFS (recurrence-free survival). The stimulated Tg level was associated with shorter RFS on multivariate analysis (>7.3 vs. ≤7.3 ng/mL; hazard ratio, 4.226; 95% confidence interval, 2.226−8.022; p < 0.001). Although patients with skip metastases tended to have a small primary tumor and lower burden of lateral neck LN involvement, there was no association between skip metastases and RFS in PTC with pN1b. Stimulated Tg level was a strong predictor of recurrence.